We have prepared crystals of proteins which represent several of the classes of actin regulatory proteins. These include actin monomer binding proteins (profilin), actin filiment cross-linking proteins (fimbrin and fascin), proteins which dictate the length of individual filaments (severin) and proteins responsible for the localixation of filamentous actin networks (ABP-1). We plan to slove these structures to atomic resolution and use this information to produce mutants which test the proposed mechanisms of action for these proteins. These mutants will be examined for in vitro actin binding activities. Once characterized, these mutant proteins will be expressed in living cells to directly determine the relationship between structure, biochemical function and in vivo activity. These studies will provide atomic level detail about the regulation of the actin cytoskeleton. This information will have broad impact for cell biology, as the actin cytoskeleton is involved in almost every fundamental physiological process in eukaryotic cells, including cell motility, transport, cytokinesis and the establishment and maintenance of cell morphology.